Project Summary Uncontrolled inflammation is a major impediment to tissue engineering, regeneration and reconstruction of both diseased and injured tissues resulting in further tissue injury, tissue scarring and fibrosis. Stem cell activity is disrupted by persistent signals promoting inflammation, whereas specific anti-inflammatory signals enhance stem cell activity. In successful regeneration, mesenchymal stem cells assume an anti-inflammatory phenotype. Specialized Proresolving lipid Mediators (SPMS), including Lipoxin A4 (LXA4), attenuate the dental stem cell inflammatory response. Resolution of inflammation is an active biochemical and metabolic process, not merely a passive termination of inflammation, mediated by SPMs. SPMs activate wound healing with tissue regeneration instead of fibrosis and scarring and directly improve bone healing and regeneration, including in periodontitis. Human periodontal ligament stem cells release SPMs, including lipoxin, to regulate immunomodulatory and pro-healing properties. Characterizing the biomimetic properties of SPMs in humans is hampered by a lack of suitable large animal models. There is a critical need for a validated large animal regeneration model to test therapeutic potential of SPMs for translation to humans. Our goal is to determine the pathways to regeneration that control local inflammation and enhance mesenchymal stem cell differentiation into connective tissues, including bone. The Central Hypothesis is that resolution of inflammation pathways and mediators can promote regeneration of the periodontal organ (bone, cementum and periodontal ligament) by directing stem cell phenotype, proliferation and differentiation. In this application, we will use a large animal model to dissect the SPM pathways leading to periodontal ligament stem cell control of regeneration. In this proposal, we will: 1: Provide direct evidence for SPM production by Yorkshire miniature pig periodontal ligament stem cells (mpPDLSC) by determining the lipid mediator profile of mpPDLSC; 2: Determine stem cell function in miniature pig by determining mpPDLSC proliferation and response to SPMs, and determination of synthetic enzyme expression, signaling pathways and SPM receptor expression, and 3: Demonstrate SPM enhanced stem cell mediated periodontal regeneration in Miniature Pigs using SPM local delivery to enhance periodontal regeneration alone or in combination with transplanted, ex vivo expanded miniature pig stem cells. Results from these studies will advance our practical clinical knowledge of dose and delivery of lipoxins in tissue regeneration, identify potential new molecular targets, and further develop and characterize a large animal model to test novel stem cell-based strategies for translation to human oral and craniofacial tissue regeneration. The research team comprises experts in periodontal regeneration, biochemistry, and large animal models.